These studies bring together laboratories who have published innovative papers on large scale microarrays to detect antibodies to proteins, peptides, lipids and carbohydrates. Epitope spreading is an immunological hallmark of multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE), an animal model of MS. It is defined as the expansion of antigen-specific immune responses beyond those targeted in the initial immunization. The complexity of the process includes spread not only to other peptide epitopes of the same protein molecule, defined as intramolecular spreading, but to other molecules, defined as intermolecular spreading. Spreading of the immune response is not confined to peptide epitopes, but also includes immune responses to lipids and carbohydrates. If tolerization to antigen specific autoimmune responses is desirable for treatment of autoimmune disease, then one must devise practical measures to tolerize the immune system across a wide front including multiple proteins/peptides, carbohydrates and lipids. We shall tolerize animals with ongoing EAE, using key proteins/peptides, lipids and carbohydrates that are targeted by autoantibodies detected on the arrays used to study both MS and EAE. We have already shown promising clinical and pre- clinical results with tolerization to these components of the myelin sheath. Both tolerization to proteins and to lipids ameliorates paralysis in EAE. We hypothesize that in order to reduce epitope spreading it will be necessary to tolerize to potentially pathogenic autoantigenic peptides/proteins, lipids AND carbohydrates, and not simply to one of these types of chemical constituents. We will now see if tolerizing individually to each of these distinct chemical components of myelin is optimal, or if tolerization in concert to proteins/peptides AND lipids AND carbohydrates, achieves even more beneficial results. In New Aim 1 we will undertake a) structure function studies on promising lipids that are the target of the immune response in MS and EAE. b) We will analyze how these tolerogens influence epitope spreading. c) We will investigate the mechanisms of action for induction of tolerance in depth on each promising candidate that suppresses ongoing EAE in three different models of EAE. In New Aim 2 we will undertake studies on tolerization to various mannose clusters that are the target of the immune response in MS and EAE, comparing for example, (Man9)n and [(Man9)4]n in reducing disease in three models of relapsing and progressive EAE. In New Aim 3, we shall tolerize animals with ongoing EAE, using SIMULTANEOUS administration of key proteins/peptides, lipids and carbohydrates shown to be targeted by autoantibodies detectable with the arrays used to study MS and EAE. We will now see if tolerizing in concert to proteins/peptides AND lipids AND carbohydrates achieves even more beneficial results in terms of reducing relapses, improving clinical function, reducing epitope spreading in three models of EAE, than tolerizing INIDIVIDUALLY to each of these three chemical types.Antigen specific tolerance is a long sought after goal for treatment of autoimmune disease. We shall develop strategies for tolerizing to proteins, lipids and carbohydrates of the myelin sheath. This approach may lead to better therapies to treat multiple sclerosis.